U.S. patent number 9,079,868 [Application Number 13/057,065] was granted by the patent office on 2015-07-14 for selective glycosidase inhibitors and uses thereof.
This patent grant is currently assigned to Simon Fraser University. The grantee listed for this patent is Ernest John McEachern, David Jaro Vocadlo. Invention is credited to Ernest John McEachern, David Jaro Vocadlo.
United States Patent |
9,079,868 |
Vocadlo , et al. |
July 14, 2015 |
Selective glycosidase inhibitors and uses thereof
Abstract
The invention provides compounds of Formula (I) for selectively
inhibiting glycosidases, prodrugs of the compounds, and
pharmaceutical compositions including the compounds or prodrugs of
the compounds. The invention also provides methods of treating
diseases and disorders related to deficiency or overexpression of
O-GlcNAcase, accumulation or deficiency of O-GlcN Ac.
##STR00001##
Inventors: |
Vocadlo; David Jaro (Burnaby,
CA), McEachern; Ernest John (Burnaby, CA) |
Applicant: |
Name |
City |
State |
Country |
Type |
Vocadlo; David Jaro
McEachern; Ernest John |
Burnaby
Burnaby |
N/A
N/A |
CA
CA |
|
|
Assignee: |
Simon Fraser University
(Burnaby, BC, CA)
|
Family
ID: |
41609897 |
Appl.
No.: |
13/057,065 |
Filed: |
July 31, 2009 |
PCT
Filed: |
July 31, 2009 |
PCT No.: |
PCT/CA2009/001088 |
371(c)(1),(2),(4) Date: |
June 10, 2011 |
PCT
Pub. No.: |
WO2010/012107 |
PCT
Pub. Date: |
February 04, 2010 |
Prior Publication Data
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Document
Identifier |
Publication Date |
|
US 20110237631 A1 |
Sep 29, 2011 |
|
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
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61085538 |
Aug 1, 2008 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61P
37/06 (20180101); A61P 17/04 (20180101); A61P
37/08 (20180101); A61P 29/00 (20180101); A61P
43/00 (20180101); A61P 1/04 (20180101); A61K
31/428 (20130101); A61P 25/28 (20180101); A61P
9/00 (20180101); A61P 11/02 (20180101); A61P
21/00 (20180101); A61P 19/02 (20180101); A61P
9/10 (20180101); A61K 31/423 (20130101); A61P
11/06 (20180101); A61P 25/04 (20180101); A61P
11/00 (20180101); A61P 5/14 (20180101); C07D
277/60 (20130101); A61P 17/00 (20180101); A61P
25/16 (20180101); A61P 35/00 (20180101); C07D
263/52 (20130101); A61P 25/00 (20180101); A61P
25/08 (20180101); A61P 17/06 (20180101); A61P
13/12 (20180101) |
Current International
Class: |
C07D
277/60 (20060101); C07D 263/52 (20060101); A61K
31/428 (20060101); A61K 31/423 (20060101) |
References Cited
[Referenced By]
U.S. Patent Documents
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11349541 |
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03/009808 |
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Feb 2003 |
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WO |
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2012/064680 |
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May 2012 |
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WO |
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2013/000084 |
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Jan 2013 |
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WO |
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2013/000085 |
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Jan 2013 |
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WO |
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2013/000086 |
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Jan 2013 |
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WO |
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2013/025452 |
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Feb 2013 |
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WO |
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2013/169576 |
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Nov 2013 |
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WO |
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2014/032184 |
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Mar 2014 |
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2014/032185 |
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2014/032187 |
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Mar 2014 |
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WO |
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2014/032188 |
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Mar 2014 |
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WO |
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2014/067003 |
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Mar 2014 |
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WO |
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2014/105662 |
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Jul 2014 |
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WO |
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|
Primary Examiner: Coughlin; Matthew
Attorney, Agent or Firm: Greenberg Traurig, LLP Pham; Chinh
H. Xie; Fang
Parent Case Text
This application is a national phase filing under 35 U.S.C.
.sctn.371 of International Application No. PCT/CA2009/001088, filed
on Jul. 31, 2009, which is hereby incorporated by reference in its
entirety for all purposes and claims the benefit of and priority to
U.S. Provisional Application No. 61/085,538, filed Aug. 1, 2008.
Claims
What is claimed is:
1.
(3aR,4R,5R,6R,6aS)-6-(hydroxymethyl)-2-(methylamino)-4,5,6,6a-tetrahyd-
ro-3aH-cyclopenta[d]oxazole-4,5-diol, or a pharmaceutically
acceptable salt thereof.
2.
(3aR,4R,5R,6R,6aS)-2-(ethylamino)-6-(hydroxymethyl)-4,5,6,6a-tetrahydr-
o-3aH-cyclopenta[d]oxazole-4,5-diol, or a pharmaceutically
acceptable salt thereof.
3.
(3aR,4R,5R,6R,6aS)-6-(hydroxymethyl)-2-(methylamino)-4,5,6,6a-tetrahyd-
ro-3aH-cyclopenta[d]thiazole-4,5-diol, or a pharmaceutically
acceptable salt thereof.
4.
(3aR,4R,5R,6R,6aS)-2-(ethylamino)-6-(hydroxymethyl)-4,5,6,6a-tetrahydr-
o-3aH-cyclopenta[d]thiazole-4,5-diol or a pharmaceutically
acceptable salt thereof.
Description
FIELD OF THE INVENTION
This application relates to compounds which selectively inhibit
glycosidases and uses thereof.
BACKGROUND OF THE INVENTION
A wide range of cellular proteins, both nuclear and cytoplasmic,
are post-translationally modified by the addition of the
monosaccharide 2-acetamido-2-deoxy-.beta.-D-glucopyranoside
(.beta.-N-acetylglucosamine) which is attached via an O-glycosidic
linkage..sup.1 This modification is generally referred to as
O-linked N-acetylglucosamine or O-GlcNAc. The enzyme responsible
for post-translationally linking .beta.-N-acetylglucosamine
(GlcNAc) to specific serine and threonine residues of numerous
nucleocytoplasmic proteins is O-GlcNAc transferase (OGT)..sup.2-5 A
second enzyme, known as O-GlcNAcase.sup.6,7 removes this
post-translational modification to liberate proteins making the
O-GlcNAc-modification a dynamic cycle occurring several times
during the lifetime of a protein..sup.8
O-GlcNAc-modified proteins regulate a wide range of vital cellular
functions including, for example, transcription,.sup.9-12
proteasomal degradation,.sup.13 and cellular signaling..sup.14
O-GlcNAc is also found on many structural proteins..sup.15-17 For
example, it has been found on a number of cytoskeletal proteins,
including neurofilament proteins,.sup.18,19 synapsins,.sup.6,20
synapsin-specific clathrin assembly protein AP-3,.sup.7 and
ankyrinG..sup.14 O-GlcNAc modification has been found to be
abundant in the brain..sup.21,22 It has also been found on proteins
clearly implicated in the etiology of several diseases including
Alzheimer's disease (AD) and cancer.
For example, it is well established that AD and a number of related
tauopathies including Downs' syndrome, Pick's disease, Niemann-Pick
Type C disease, and amyotrophic lateral sclerosis (ALS) are
characterized, in part, by the development of neurofibrillary
tangles (NFTs). These NFTs are aggregates of paired helical
filaments (PHFs) and are composed of an abnormal form of the
cytoskeletal protein "tau". Normally tau stabilizes a key cellular
network of microtubules that is essential for distributing proteins
and nutrients within neurons. In AD patients, however, tau becomes
hyperphosphorylated, disrupting its normal functions, forming PHFs
and ultimately aggregating to form NFTs. Six isoforms of tau are
found in the human brain. In AD patients, all six isoforms of tau
are found in NFTs, and all are markedly
hyperphosphorylated..sup.23,24 Tau in healthy brain tissue bears
only 2 or 3 phosphate groups, whereas those found in the brains of
AD patients bear, on average, 8 phosphate groups..sup.25,26 A clear
parallel between NFT levels in the brains of AD patients and the
severity of dementia strongly supports a key role for tau
dysfunction in AD..sup.27,28 The precise causes of this
hyperphosphorylation of tau remain elusive. Accordingly,
considerable effort has been dedicated toward: a) elucidating the
molecular physiological basis of tau hyperphosphorylation;.sup.29
and b) identifying strategies that could limit tau
hyperphosphorylation in the hope that these might halt, or even
reverse, the progression of Alzheimer's disease.sup.30-33 Thus far,
several lines of evidence suggest that up-regulation of a number of
kinases may be involved in hyperphosphorylation of
tau,.sup.21,34,35 although very recently, an alternative basis for
this hyperphosphorylation has been advanced..sup.21
In particular, it has recently emerged that phosphate levels of tau
are regulated by the levels of O-GlcNAc on tau. The presence of
O-GlcNAc on tau has stimulated studies that correlate O-GlcNAc
levels with tau phosphorylation levels. The recent interest in this
field stems from the observation that O-GlcNAc modification has
been found to occur on many proteins at amino acid residues that
are also known to be phosphorylated..sup.36-38 Consistent with this
observation, it has been found that increases in phosphorylation
levels result in decreased O-GlcNAc levels and conversely,
increased O-GlcNAc levels correlate with decreased phosphorylation
levels..sup.39 This reciprocal relationship between O-GlcNAc and
phosphorylation has been termed the "Yin-Yang hypothesis".sup.40
and has gained strong biochemical support by the recent discovery
that the enzyme OGT.sup.4 forms a functional complex with
phosphatases that act to remove phosphate groups from
proteins..sup.41 Like phosphorylation, O-GlcNAc is a dynamic
modification that can be removed and reinstalled several times
during the lifespan of a protein. Suggestively, the gene encoding
O-GlcNAcase has been mapped to a chromosomal locus that is linked
to AD..sup.7,42 Hyperphosphorylated tau in human AD brains has
markedly lower levels of O-GlcNAc than are found in healthy human
brains..sup.21 Very recently, it has been shown that O-GlcNAc
levels of soluble tau protein from human brains affected with AD
are markedly lower than those from healthy brain..sup.21
Furthermore, PHF from diseased brain was suggested to lack
completely any O-GlcNAc modification whatsoever..sup.21 The
molecular basis of this hypoglycosylation of tau is not known,
although it may stem from increased activity of kinases and/or
dysfunction of one of the enzymes involved in processing O-GlcNAc.
Supporting this latter view, in both PC-12 neuronal cells and in
brain tissue sections from mice, a nonselective
N-acetylglucosamindase inhibitor was used to increase tau O-GlcNAc
levels, whereupon it was observed that phosphorylation levels
decreased..sup.21 The implication of these collective results is
that by maintaining healthy O-GlcNAc levels in AD patients, such as
by inhibiting the action of O-GlcNAcase, one should be able to
block hyperphosphorylation of tau and all of the associated effects
of tau hyperphosphorylation, including the formation of NFTs and
downstream effects. However, because the proper functioning of the
.beta.-hexosaminidases is critical, any potential therapeutic
intervention for the treatment of AD that blocks the action of
O-GlcNAcase would have to avoid the concomitant inhibition of both
hexosaminidases A and B.
Neurons do not store glucose and therefore the brain relies on
glucose supplied by blood to maintain its essential metabolic
functions. Notably, it has been shown that within brain, glucose
uptake and metabolism decreases with aging..sup.43 Within the
brains of AD patients marked decreases in glucose utilization occur
and are thought to be a potential cause of
neurodegeneration..sup.44 The basis for this decreased glucose
supply in AD brain.sup.45-47 is thought to stem from any of
decreased glucose transport,.sup.48,49 impaired insulin
signaling,.sup.50,51 and decreased blood flow..sup.52
In light of this impaired glucose metabolism, it is worth noting
that of all glucose entering into cells, 2-5% is shunted into the
hexosamine biosynthetic pathway, thereby regulating cellular
concentrations of the end product of this pathway, uridine
diphosphate-N-acetylglucosamine (UDP-GlcNAc)..sup.53 UDP-GlcNAc is
a substrate of the nucleocytoplasmic enzyme O-GlcNAc transferase
(OGT),.sup.2-5 which acts to post-translationally add GlcNAc to
specific serine and threonine residues of numerous
nucleocytoplasmic proteins. OGT recognizes many of its
substrates.sup.54,55 and binding partners.sup.41,56 through its
tetratricopeptide repeat (TPR) domains..sup.57,58 As described
above, O-GlcNAcase.sup.6,7 removes this post-translational
modification to liberate proteins making the O-GlcNAc-modification
a dynamic cycle occurring several times during the lifetime of a
protein..sup.8 O-GlcNAc has been found in several proteins on known
phosphorylation sites,.sup.10,37,38,59 including tau and
neurofilaments..sup.60 Additionally, OGT shows unusual kin
behaviour making it exquisitely sensitive to intracellular
UDP-GlcNAc substrate concentrations and therefore glucose
supply..sup.41
Consistent with the known properties of the hexosamine biosynthetic
pathway, the enzymatic properties of OGT, and the reciprocal
relationship between O-GlcNAc and phosphorylation, it has been
shown that decreased glucose availability in brain leads to tau
hyperphosphorylation..sup.44 Therefore the gradual impairment of
glucose transport and metabolism, whatever its causes, leads to
decreased O-GlcNAc and hyperphosphorylation of tau (and other
proteins). Accordingly, the inhibition of O-GlcNAcase should
compensate for the age related impairment of glucose metabolism
within the brains of health individuals as well as patients
suffering from AD or related neurodegenerative diseases.
These results suggest that a malfunction in the mechanisms
regulating tau O-GlcNAc levels may be vitally important in the
formation of NFTs and associated neurodegeneration. Good support
for blocking tau hyperphosphorylation as a therapeutically useful
intervention.sup.61 comes from recent studies showing that when
transgenic mice harbouring human tau are treated with kinase
inhibitors, they do not develop typical motor defects.sup.33 and,
in another case,.sup.32 show decreased levels of insoluble tau.
These studies provide a clear link between lowering tau
phosphorylation levels and alleviating AD-like behavioural symptoms
in a murine model of this disease. Indeed, pharmacological
modulation of tau hyperphosphorylation is widely recognized as a
valid therapeutic strategy for treating AD and other
neurodegenerative disorders..sup.62
Recent studies.sup.63 support the therapeutic potential of
small-molecule O-GlcNAcase inhibitors to limit tau
hyperphosphorylation for treatment of AD and related tauopathies.
Specifically, the O-GlcNAcase inhibitor thiamet-G has been
implicated in the reduction of tau phosphorylation in cultured
PC-12 cells at pathologically relevant sites..sup.63 Moreover, oral
administration of thiamet-G to healthy Sprague-Dawley rats has been
implicated in reduced phosphorylation of tau at Thr231, Ser396 and
Ser422 in both rat cortex and hippocampus..sup.63
There is also a large body of evidence indicating that increased
levels of O-GlcNAc protein modification provides protection against
pathogenic effects of stress in cardiac tissue, including stress
caused by ischemia, hemorrhage, hypervolemic shock, and calcium
paradox. For example, activation of the hexosamine biosynthetic
pathway (HBP) by administration of glucosamine has been
demonstrated to exert a protective effect in animals models of
ischemia/reperfusion.sup.64-70 trauma hemorrhage,.sup.71-73
hypervolemic shock,.sup.74 and calcium paradox..sup.64,75 Moreover,
strong evidence indicates that these cardioprotective effects are
mediated by elevated levels of protein O-GlcNAc
modification..sup.64,65,67,70,72,75-78 There is also evidence that
the O-GlcNAc modification plays a role in a variety of
neurodegenerative diseases, including Parkinson's disease and
Huntington's disease..sup.79
Humans have three genes encoding enzymes that cleave terminal
.beta.-N-acetyl-glucosamine residues from glycoconjugates. The
first of these encodes the enzyme O-glycoprotein
2-acetamido-2-deoxy-.beta.-D-glucopyranosidase (O-GlcNAcase).
O-GlcNAcase is a member of family 84 of glycoside hydrolases that
includes enzymes from organisms as diverse as prokaryotic pathogens
to humans (for the family classification of glycoside hydrolases
see Coutinho, P. M. & Henrissat, B. (1999) Carbohydrate-Active
Enzymes server at URL: http://afmb.cnrs-mrs.fr/CAZY/..sup.27,28
O-GlcNAcase acts to hydrolyse O-GlcNAc off of serine and threonine
residues of post-translationally modified proteins..sup.1,6,7,80,81
Consistent with the presence of O-GlcNAc on many intracellular
proteins, the enzyme O-GlcNAcase appears to have a role in the
etiology of several diseases including type II diabetes,.sup.14,82
AD.sup.16,21,83 and cancer..sup.22,84 Although O-GlcNAcase was
likely isolated earlier on,.sup.18,19 about 20 years elapsed before
its biochemical role in acting to cleave O-GlcNAc from serine and
threonine residues of proteins was understood..sup.6 More recently
O-GlcNAcase has been cloned,.sup.7 partially characterized,.sup.20
and suggested to have additional activity as a histone
acetyltransferase..sup.20 However, little was known about the
catalytic mechanism of this enzyme.
The other two genes, HEXA and HEXB, encode enzymes catalyzing the
hydrolytic cleavage of terminal .beta.-N-acetylglucosamine residues
from glycoconjugates. The gene products of HEXA and HEXB
predominantly yield two dimeric isozymes, hexosaminidase A and
hexosaminidase B, respectively. Hexosaminidase A (.alpha..beta.), a
heterodimeric isozyme, is composed of an .alpha.- and a
.beta.-subunit. Hexosaminidase B (.beta..beta.), a homodimeric
isozyme, is composed of two .beta.-subunits. The two subunits,
.alpha.- and .beta.-, bear a high level of sequence identity. Both
of these enzymes are classified as members of family 20 of
glycoside hydrolases and are normally localized within lysosomes.
The proper functioning of these lysosomal .beta.-hexosaminidases is
critical for human development, a fact that is underscored by the
tragic genetic illnesses, Tay-Sach's and Sandhoff diseases which
stem from a dysfunction in, respectively, hexosaminidase A and
hexosaminidase B..sup.85 These enzymatic deficiencies cause an
accumulation of glycolipids and glycoconjugates in the lysosomes
resulting in neurological impairment and deformation. The
deleterious effects of accumulation of gangliosides at the
organismal level are still being uncovered..sup.86
As a result of the biological importance of these
.beta.-N-acetyl-glucosaminidases, small molecule inhibitors of
glycosidases.sup.87-90 have received a great deal of
attention,.sup.91 both as tools for elucidating the role of these
enzymes in biological processes and in developing potential
therapeutic applications. The control of glycosidase function using
small molecules offers several advantages over genetic knockout
studies including the ability to rapidly vary doses or to entirely
withdraw treatment.
However, a major challenge in developing inhibitors for blocking
the function of mammalian glycosidases, including O-GlthAcase, is
the large number of functionally related enzymes present in tissues
of higher eukaryotes. Accordingly, the use of non-selective
inhibitors in studying the cellular and organismal physiological
role of one particular enzyme is complicated because complex
phenotypes arise from the concomitant inhibition of such
functionally related enzymes. In the case of
.beta.-N-acetylglucosaminidases, existing compounds that act to
block O-GlcNAcase function are non-specific and act potently to
inhibit the lysosomal .beta.-hexosaminidases.
A few of the better characterized inhibitors of
.beta.-N-acetyl-glucosaminidases which have been used in studies of
O-GlcNAc post-translational modification within both cells and
tissues are streptozotocin (STZ),
2'-methyl-.alpha.-D-glucopyrano-[2,1-d]-.DELTA.2'-thiazoline
(NAG-thiazoline) and
O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino
N-phenylcarbamate (PUGNAc)..sup.14,92-95
STZ has long been used as a diabetogenic compound because it has a
particularly detrimental effect on .beta.-islet cells..sup.96 STZ
exerts its cytotoxic effects through both the alkylation of
cellular DNA.sup.96,97 as well as the generation of radical species
including nitric oxide..sup.98 The resulting DNA strand breakage
promotes the activation of poly(ADP-ribose) polymerase
(PARP).sup.99 with the net effect of depleting cellular NAD+ levels
and, ultimately, leading to cell death..sup.100,101 Other
investigators have proposed instead that STZ toxicity is a
consequence of the irreversible inhibition of O-GlcNAcase, which is
highly expressed within .beta.-islet cells..sup.92,102 This
hypothesis has, however, been brought into question by two
independent research groups..sup.103,104 Because cellular O-GlcNAc
levels on proteins increase in response to many forms of cellular
stress.sup.105 it seems possible that STZ results in increased
O-GlcNAc-modification levels on proteins by inducing cellular
stress rather than through any specific and direct action on
O-GlcNAcase. Indeed, Hanover and coworkers have shown that STZ
functions as a poor and somewhat selective inhibitor of
O-GlcNAcase.sup.106 and although it has been proposed by others
that STZ acts to irreversibly inhibit O-GlcNAcase,.sup.107 there
has been no clear demonstration of this mode of action. Recently,
it has been shown that STZ does not irreversibly inhibit
O-GlcNAcase..sup.108
NAG-thiazoline has been found to be a potent inhibitor of family 20
hexosaminidases,.sup.90,109 and more recently, the family 84
O-GlcNAcases..sup.108 Despite its potency, a downside to using
NAG-thiazoline in a complex biological context is that it lacks
selectivity and therefore perturbs multiple cellular processes.
PUGNAc is another compound that suffers from the same problem of
lack of selectivity, yet has enjoyed use as an inhibitor of both
human O-GlcNAcase.sup.6,110 and the family 20 human
.beta.-hexosaminidases..sup.111 This molecule, developed by Vasella
and coworkers, was found to be a potent competitive inhibitor of
the .beta.-N-acetyl-glucosaminidases from Canavalia ensiformis,
Mucor rouxii, and the .beta.-hexosaminidase from bovine
kidney..sup.88 It has been demonstrated that administration of
PUGNAc in a rat model of trauma hemorrhage decreases circulating
levels of the pro-inflammatory cytokines TNF-.alpha. and
IL-6..sup.112 It has also been shown that administration of PUGNAc
in a cell-based model of lymphocyte activation decreases production
of the cytokine IL-2..sup.113 Recent studies have indicated that
PUGNAc can be used in an animal model to reduce myocardial infarct
size after left coronary artery occlusions..sup.114 Of particular
significance is the fact that elevation of O-GlcNAc levels by
administration of PUGNAc, an inhibitor of O-GlcNAcase, in a rat
model of trauma hemorrhage improves cardiac function..sup.112,115
In addition, elevation of O-GlcNAc levels by treatment with PUGNAc
in a cellular model of ischemia/reperfusion injury using neonatal
rat ventricular myocytes improved cell viability and reduced
necrosis and apoptosis compared to untreated cells..sup.116
More recently, it has been suggested that the selective O-GlcNAcase
inhibitor NButGT exhibits protective activity in cell-based models
of ischemia/reperfusion and cellular stresses, including oxidative
stress..sup.117 This evidence suggests the use of O-GlcNAcase
inhibitors to elevate protein O-GlcNAc levels and thereby prevent
the pathogenic effects of stress in cardiac tissue.
International patent applications PCT/CA2006/000300, filed 1 Mar.
2006, published under No. WO 2006/092049 on 8 Sep. 2006, and
PCT/CA2007/001554, filed 31 Aug. 2007, published under No. WO
2008/025170 on 6 Mar. 2008, which are hereby incorporated by
reference, describe selective inhibitors of O-GlcNAcase
SUMMARY OF THE INVENTION
The invention provides, in part, compounds for selectively
inhibiting glycosidases, prodrugs of the compounds, uses of the
compounds and the prodrugs, pharmaceutical compositions including
the compounds or prodrugs of the compounds, and methods of treating
diseases and disorders related to deficiency or overexpression of
O-GlcNAcase, and/or accumulation or deficiency of O-GlcNAc.
In one aspect, the invention provides a compound of Formula (I) or
a pharmaceutically acceptable salt thereof:
##STR00002##
where each R.sup.1 may be independently a non-interfering
substituent; X may be O, S, or NR.sup.3; R.sup.2 is NR.sup.3.sub.2;
where each R.sup.3 may be optionally independently a
non-interfering substituent, with the proviso that when X is O and
each R.sup.1 is H or C(O)CH.sub.3, R.sup.2 excludes
N(CH.sub.3).sub.2.
In alternative embodiments, each R.sup.1 may be connected to
another R.sup.1 to form an additional ring structure.
In alternative embodiments, the non-interfering substituent may be
alkyl, alkenyl, alkynyl, aryl, arylalkyl, arylalkenyl, or
arylalkynyl, or may include one or more heteroatoms selected from
P, O, S, N, F, Cl, Br, I, or B. The non-interfering substituent may
be optionally substituted.
In alternative embodiments, the compound may be a prodrug; the
compound may selectively inhibit an O-glycoprotein
2-acetamido-2-deoxy-.beta.-D-glucopyranosidase (O-GlcNAcase); the
compound may selectively bind an O-GlcNAcase (e.g., a mammalian
O-GlcNAcase); the compound may selectively inhibit the cleavage of
a 2-acetamido-2-deoxy-.beta.-D-glucopyranoside (O-GlcNAc); the
compound may not substantially inhibit a mammalian
.beta.-hexosaminidase.
In alternative aspects, the invention provides a pharmaceutical
composition including a compound according to the invention, in
combination with a pharmaceutically acceptable carrier.
In alternative aspects, the invention provides methods of
selectively inhibiting an O-GlcNAcase, or of inhibiting an
O-GlcNAcase in a subject in need thereof, or of increasing the
level of O-GlcNAc, or of treating a neurodegenerative disease, a
tauopathy, cancer or stress, in a subject in need thereof, by
administering to the subject an effective amount of a compound of
Formula (I) or a pharmaceutically acceptable salt thereof:
##STR00003##
wherein each R.sup.1 may be independently a non-interfering
substituent; X may be O, S, or NR.sup.3; R.sup.2 may be
NR.sup.3.sub.2; where each R.sup.3 may be optionally independently
a non-interfering substituent. The condition may be Alzheimer's
disease, Amyotrophic lateral sclerosis (ALS), Amyotrophic lateral
sclerosis with cognitive impairment (ALSci), Argyrophilic grain
dementia, Bluit disease, Corticobasal degeneration (CBD), Dementia
pugilistica, Diffuse neurofibrillary tangles with calcification,
Down's syndrome, Familial British dementia, Familial Danish
dementia, Frontotemporal dementia with parkinsonism linked to
chromosome 17 (FTDP-17), Gerstmann-Straussler-Scheinker disease,
Guadeloupean parkinsonism, Hallevorden-Spatz disease
(neurodegeneration with brain iron accumulation type 1), Multiple
system atrophy, Myotonic dystrophy, Niemann-Pick disease (type C),
Pallido-ponto-nigral degeneration, Parkinsonism-dementia complex of
Guam, Pick's disease (PiD), Post-encephalitic parkinsonism (PEP),
Prion diseases (including Creutzfeldt-Jakob Disease (CJD), Variant
Creutzfeldt-Jakob Disease (vCJD), Fatal Familial Insomnia, and
Kuru), Progressive supercortical gliosis, Progressive supranuclear
palsy (PSP), Richardson's syndrome, Subacute sclerosing
panencephalitis, Tangle-only dementia, Huntington's disease, or
Parkinson's disease. The stress may be a cardiac disorder, e.g.,
ischemia; hemorrhage; hypovolemic shock; myocardial infarction; an
interventional cardiology procedure; cardiac bypass surgery;
fibrinolytic therapy; angioplasty; or stent placement.
In alternative aspects, the invention provides a method of treating
an O-GlcNAcase-mediated condition that excludes a neurodegenerative
disease, a tauopathy, cancer or stress, in a subject in need
thereof, by administering to the subject an effective amount of a
compound of Formula (I) or a pharmaceutically acceptable salt
thereof:
##STR00004##
wherein each R.sup.1 may be independently a non-interfering
substituent; X may be O, S, or NR.sup.3; R.sup.2 may be
NR.sup.3.sub.2; where each R.sup.3 may be optionally independently
a non-interfering substituent. In some embodiments, the condition
may be inflammatory or allergic diseases such as asthma, allergic
rhinitis, hypersensitivity lung diseases, hypersensitivity
pneumonitis, eosinophilic pneumonias, delayed-type
hypersensitivity, atherosclerosis, interstitial lung disease (ILD)
(e.g., idiopathic pulmonary fibrosis, or ILD associated with
rheumatoid arthritis, systemic lupus erythematosus, ankylosing
spondylitis, systemic sclerosis, Sjogren's syndrome, polymyositis
or dermatomyositis); systemic anaphylaxis or hypersensitivity
responses, drug allergies, insect sting allergies; autoimmune
diseases, such as rheumatoid arthritis, psoriatic arthritis,
multiple sclerosis, systemic lupus erythematosus, myastenia gravis,
glomerulonephritis, autoimmune thyroiditis, graft rejection,
including allograft rejection or graft-versus-host disease;
inflammatory bowel diseases, such as Crohn's disease and ulcerative
colitis; spondyloarthropathies; scleroderma; psoriasis (including
T-cell mediated psoriasis) and inflammatory dermatoses such as
dermatitis, eczema, atopic dermatitis, allergic contact dermatitis,
urticaria; vasculitis (e.g., necrotizing, cutaneous, and
hypersensitivity vasculitis); eosinphilic myotis, and eosiniphilic
fasciitis; graft rejection, in particular but not limited to solid
organ transplants, such as heart, lung, liver, kidney, and pancreas
transplants (e.g. kidney and lung allografts); epilepsy; pain;
stroke, e.g., neuroprotection following a stroke.
In alternative embodiments, X may be O; R.sup.1 may be H or
C(O)CH.sub.3. The administering may increase the level of O-GlcNAc
in the subject. The subject may be a human.
In alternative aspects, the invention provides use of a compound of
an effective amount of a compound of Formula (I) or a
pharmaceutically acceptable salt thereof:
##STR00005##
where each R.sup.1 may be independently a non-interfering
substituent; X may be O, S, or NR.sup.3; R.sup.2 may be
NR.sup.3.sub.2; where each R.sup.3 may be optionally independently
a non-interfering substituent, in the preparation of a medicament.
The medicament may be for selectively inhibiting an O-GlcNAcase,
for increasing the level of O-GlcNAc, for treating a condition
modulated by an O-GlcNAcase, for treating a neurodegenerative
disease, a tauopathy, a cancer, or stress.
In alternative aspects, the invention provides a method for
screening for a selective inhibitor of an O-GlcNAcase, by a)
contacting a first sample with a test compound; b) contacting a
second sample with a compound of Formula (I)
##STR00006##
where each R.sup.1 may be independently a non-interfering
substituent; X may be O, S, or NR.sup.3; R.sup.2 may be
NR.sup.3.sub.2; where each R.sup.3 may be optionally independently
a non-interfering substituent, c) determining the level of
inhibition of the O-GlcNAcase in the first and second samples,
where the test compound is a selective inhibitor of a O-GlcNAcase
if the test compound exhibits the same or greater inhibition of the
O-GlcNAcase when compared to the compound of Formula (I).
This summary of the invention does not necessarily describe all
features of the invention.
DETAILED DESCRIPTION
The invention provides, in part, novel compounds that are capable
of inhibiting an O-glycoprotein
2-acetamido-2-deoxy-.beta.-D-glucopyranosidase (O-GlcNAcase). In
some embodiments, the O-GlcNAcase is a mammalian O-GlcNAcase, such
as a rat, mouse or human O-GlcNAcase. In some embodiments, the
.beta.-hexosaminidase is a mammalian .beta.-hexosaminidase, such as
a rat, mouse or human .beta.-hexosaminidase.
In some embodiments, compounds according to the invention exhibit a
surprising and unexpected selectivity in inhibiting an O-GlcNAcase.
In some embodiments, the compounds according to the invention are
surprisingly more selective for an O-GlcNAcase over a
.beta.-hexosaminidase. In some embodiments, the compounds
selectively inhibit the activity of a mammalian O-GlcNAcase over a
mammalian .beta.-hexosaminidase. In some embodiments, a selective
inhibitor of an O-GlcNAcase does not substantially inhibit a
.beta.-hexosaminidase. A compound that "selectively" inhibits an
O-GlcNAcase is a compound that inhibits the activity or biological
function of an O-GlcNAcase, but does not substantially inhibit the
activity or biological function of a .beta.-hexosaminidase. For
example, in some embodiments, a selective inhibitor of an
O-GlcNAcase selectively inhibits the cleavage of
2-acetamido-2-deoxy-.beta.-D-glucopyranoside (O-GlcNAc) from
polypeptides. In some embodiments, a selective inhibitor of an
O-GlcNAcase selectively binds to an O-GlcNAcase. In some
embodiments, a selective inhibitor of an O-GlcNAcase inhibits
hyperphosphorylation of a tau protein and/or inhibits formations of
NFTs. By "inhibits," "inhibition" or "inhibiting" means a decrease
by any value between 10% and 90%, or of any integer value between
30% and 60%, or over 100%, or a decrease by 1-fold, 2-fold, 5-fold,
10-fold or more. It is to be understood that the inhibiting does
not require full inhibition. In some embodiments, a selective
inhibitor of an O-GlcNAcase elevates or enhances O-GlcNAc levels
e.g., O-GlcNAc-modified polypeptide or protein levels, in cells,
tissues, or organs (e.g., in brain, muscle, or heart (cardiac)
tissue) and in animals. By "elevating" or "enhancing" is meant an
increase by any value between 10% and 90%, or of any integer value
between 30% and 60%, or over 100%, or an increase by 1-fold,
2-fold, 5-fold, 10-fold, 15-fold, 25-fold, 50-fold, 100-fold or
more. In some embodiments, a selective inhibitor of an O-GlcNAcase
exhibits a selectivity ratio, as described herein, in the range 100
to 100000, or in the range 1000 to 100000, or at least 100, 200,
500, 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 6000,
7000, 10,000, 25,000, 50,000, 75,000, or any value within or about
the described range.
The compounds of the present invention elevate O-GlcNAc levels on
O-GlcNAc-modified polypeptides or proteins in vivo specifically via
interaction with an O-GlcNAcase enzyme, and are effective in
treating conditions which require or respond to inhibition of
O-GlcNAcase activity.
In some embodiments, the compounds of the present invention are
useful as agents that produce a decrease in tau phosphorylation and
NFT formation. In some embodiments, the compounds are therefore
useful to treat Alzheimer's disease and related tauopathies. In
some embodiments, the compounds are thus capable of treating
Alzheimer's disease and related tauopathies by lowering tau
phosphorylation and reducing NFT formation as a result of
increasing tau O-GlcNAc levels. In some embodiments, the compounds
produce an increase in levels of O-GlcNAc modification on
O-GlcNAc-modified polypeptides or proteins, and are therefore
useful for treatment of disorders responsive to such increases in
O-GlcNAc modification; these disorders include without limitation
neurodegenerative, inflammatory, cardiovascular, and
immunoregulatory diseases. In some embodiments, the compounds are
also useful as a result of other biological activities related to
their ability to inhibit the activity of glycosidase enzymes. In
alternative embodiments, the compounds of the invention are
valuable tools in studying the physiological role of O-GlcNAc at
the cellular and organismal level.
In alternative embodiments, the invention provides methods of
enhancing or elevating levels of protein O-GlcNAc modification in
animal subjects, such as, veterinary and human subjects. In
alternative embodiments, the invention provides methods of
selectively inhibiting an O-GlcNAcase enzyme in animal subjects,
such as, veterinary and human subjects. In alternative embodiments,
the invention provides methods of inhibiting phosphorylation of tau
polypeptides, or inhibiting formation of NFTs, in animal subjects,
such as, veterinary and human subjects.
In specific embodiments, the invention provides compounds described
generally by Formula (I) and the salts, prodrugs, and
stereoisomeric forms thereof:
##STR00007##
As set forth in Formula (I): each R.sup.1 can be independently a
non-interfering substituent; X can be O, S, or NR.sup.3; R.sup.2
can be NR.sup.3.sub.2; where each R.sup.3 may be optionally
independently a non-interfering substituent. In some embodiments,
each R.sup.1 may be connected to another R.sup.1 to form an
additional ring structure.
In the above Formula (I), each optionally substituted moiety may be
substituted with one or more non-interfering substituents. For
example, each optionally substituted moiety may be substituted with
one or more inorganic substituents; phosphoryl; halo; .dbd.O;
.dbd.NR.sup.4; OR; C.sub.1-10 alkyl or C.sub.2-10 alkenyl
optionally containing one or more P, N, O, S, N, F, Cl, Br, I, or
B, and optionally substituted with halo; CN; optionally substituted
carbonyl; NR.sup.4.sub.2; C.dbd.NR.sup.4; an optionally substituted
carbocyclic or heterocyclic ring; or an optionally substituted aryl
or heteroaryl. R.sup.4 may be alkyl, branched alkyl, cycloalkyl,
aryl, or heteroaryl.
In some embodiments, R.sup.1 as set forth in Formula (I), may be
either hydrogen or a substituent that includes 1-20 atoms that are
other than hydrogen. In some embodiments, R.sup.1 may be H, alkyl,
or C(O)R.sup.4, where R.sup.4 may be alkyl, branched alkyl,
cycloalkyl, aryl, or heteroaryl. In some embodiments, R.sup.1 may
be H or C(O)CH.sub.3.
In some embodiments, R.sup.2 as set forth in Formula (I), may be
optionally substituted NR.sup.5.sub.2, where R.sup.5 may be H,
alkyl, branched alkyl, cycloalkyl, aryl, or heteroaryl. In some
embodiments, R.sup.2 may be N(CH.sub.3).sub.2.
In specific embodiments of the invention, compounds according to
Formula (I) include the compounds described in Table 1.
TABLE-US-00001 TABLE 1 Compound Name Structure 1
(3aS,4R,5R,6S,6aS)-6- (acetoxymethyl)-2- (dimethylamino)-
4,5,6,6a-tetrahydro-3aH- cyclopenta[d]oxazole- 4,5-diyl diacetate
##STR00008## 2 (3aR,4R,5R,6R,6aS)-2- (dimethylamino)-6-
(hydroxymethyl)- 4,5,6,6a-tetrahydro-3aH- cyclopenta[d]oxazole-
4,5-diol ##STR00009##
In alternative embodiments of the invention, compounds according to
Formula (I) include one or more of the compounds described in Table
2.
TABLE-US-00002 TABLE 2 Com- pound Name Structure 3
(3aR,4R,5R,6R,6aS)- 2-amino- 6-(hydroxymethyl)- 4,5,6,6a-
tetrahydro-3aH- cyclopenta[d]oxazole- 4,5-diol ##STR00010## 4
(3aR,4R,5R,6R,6aS)-6- (hydroxymethyl)-2- (methylamino)- 4,5,6,6a-
tetrahydro-3aH- cyclopenta[d]oxazole- 4,5-diol ##STR00011## 5
(3aR,4R,5R,6R,6aS)-2- (ethylamino)-6- (hydroxymethyl)- 4,5,6,6a-
tetrahydro-3aH- cyclopenta[d]oxazole- 4,5-diol ##STR00012## 6
(3aR,4R,5R,6R,6aS)-6- (hydroxymethyl)-2- (propylamino)- 4,5,6,6a-
tetrahydro-3aH- cyclopenta[d]oxazole- 4,5-diol ##STR00013## 7
(3aR,4R,5R,6R,6aR)-2- (butylamino)-6- (hydroxymethyl)- 4,5,6,6a-
tetrahydro-3aH- cyclopenta[d]oxazole- 4,5-diol ##STR00014## 8
(3aR,4R,5R,6R,6aS)-2- (allylamino)-6- (hydroxymethyl)- 4,5,6,6a-
tetrahydro-3 aH- cyclopenta[d]oxazole- 4,5-diol ##STR00015## 9
(3aR,4R,5R,6R,6aS)-2- (ethyl(methyl)amino)-6- (hydroxymethyl)-
4,5,6,6a- tetrahydro-3aH- cyclopenta[d]oxazole- 4,5-diol
##STR00016## 10 (3aR,4R,5R,6R,6aS)-2- (diethylamino)-6-
(hydroxymethyl)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]oxazole-
4,5-diol ##STR00017## 11 (3aR,4R,5R,6R,6aS)-6- (hydroxymethyl)-2-
(pyrrolidin-1-yl)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]oxazole-
4,5-diol ##STR00018## 12 (3aR,4R,5R,6R,6aS)- 2-amino-
6-(hydroxymethyl)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]thiazole-
4,5-diol ##STR00019## 13 (3aR,4R,5R,6R,6aS)-6- (hydroxymethyl)-2-
(methylamino)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]thiazole-
4,5-diol ##STR00020## 14 (3aR,4R,5R,6R,6aS)-2- (ethylamino)-6-
(hydroxymethyl)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]thiazole-
4,5-diol ##STR00021## 15 (3aR,4R,5R,6R,6aS)-6- (hydroxymethyl)-2-
(propylamino)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]thiazole-
4,5-diol ##STR00022## 16 (3aR,4R,5R,6R,6aS)-2- (butylamino)-6-
(hydroxymethyl)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]thiazole-
4,5-diol ##STR00023## 17 (3aR,4R,5R,6R,6aS)-2- (allylamino)-6-
(hydroxymethyl)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]thiazole-
4,5-diol ##STR00024## 18 (3aR,4R,5R,6R,6aR)-2- (dimethylamino)-6-
(hydroxymethyl)- 4,5,6,6a- tetrahydro-3aH- cyclopenta[d]thiazole-
4,5-diol ##STR00025## 19 (3aR,4R,5R,6R,6aS)-2- (ethyl(methyl)
amino)-6- (hydroxymethyl)- 4,5,6,6a- tetrahydro-3aH-
cyclopenta[d]thiazole- 4,5-diol ##STR00026## 20
(3aR,4R,5R,6R,6aS)-2- (diethylamino)-6- (hydroxymethyl)- 4,5,6,6a-
tetrahydro-3aH- cyclopenta[d]thiazole- 4,5-diol ##STR00027## 21
(3aR,4R,5R,6R,6aS)-6- (hydroxymethyl)-2- (pyrrolidin-1-yl)-
4,5,6,6a- tetrahydro-3aH- cyclopenta[d]thiazole- 4,5-diol
##STR00028##
In alternative embodiments of the invention, one or more of the
compounds described in Table 1 are specifically excluded from the
compounds described in Formula (I). In alternative embodiments of
the invention, specific stereoisomers or enantiomers of one or more
of the compounds described in Table 1 are specifically excluded
from the compounds described in Formula (I). In alternative
embodiments of the invention, specific precursors of one or more of
the compounds described in Table 1 are specifically excluded from
the compounds described in Formula (I).
In alternative embodiments, when X is O and each R.sup.1 is H or
C(O)CH.sub.3, R.sup.2 excludes N(CH.sub.3).sub.2.
As will be appreciated by a person skilled in the art, Formula (I)
above may also be represented alternatively as follows:
##STR00029##
As used herein the singular forms "a", "and", and "the" include
plural referents unless the context clearly dictates otherwise. For
example, "a compound" refers to one or more of such compounds,
while "the enzyme" includes a particular enzyme as well as other
family members and equivalents thereof as known to those skilled in
the art.
Throughout this application, it is contemplated that the term
"compound" or "compounds" refers to the compounds discussed herein
and includes precursors and derivatives of the compounds, including
acyl-protected derivatives, and pharmaceutically acceptable salts
of the compounds, precursors, and derivatives. The invention also
includes prodrugs of the compounds, pharmaceutical compositions
including the compounds and a pharmaceutically acceptable carrier,
and pharmaceutical compositions including prodrugs of the compounds
and a pharmaceutically acceptable carrier.
In some embodiments, all of the compounds of the invention contain
at least one chiral center. In some embodiments, the formulations,
preparation, and compositions including compounds according to the
invention include mixtures of stereoisomers, individual
stereoisomers, and enantiomeric mixtures, and mixtures of multiple
stereoisomers. In general, the compound may be supplied in any
desired degree of chiral purity.
In general, a "non-interfering substituent" is a substituent whose
presence does not destroy the ability of the compound of Formula
(I) to modulate the activity of the O-GlcNAcase enzyme.
Specifically, the presence of the substituent does not destroy the
effectiveness of the compound as a modulator of the activity of the
O-GlcNAcase enzyme.
Suitable non-interfering substituents include: H, alkyl
(C.sub.1-10, alkenyl (C.sub.2-10, alkynyl (C.sub.2-10), aryl (5-12
members), arylalkyl, arylalkenyl, or arylalkynyl, each of which may
optionally contain one or more heteroatoms selected from O, S, P,
N, F, Cl, Br, I, or B, and each of which may be further
substituted, for example, by .dbd.O; or optionally substituted
forms of acyl, arylacyl, alkyl- alkenyl-, alkynyl- or arylsulfonyl
and forms thereof which contain heteroatoms in the alkyl, alkenyl,
alkynyl or aryl moieties. Other noninterfering substituents include
.dbd.O, .dbd.NR, halo, CN, CF.sub.3, CHF.sub.2, NO.sub.2, OR, SR,
NR.sub.2, N.sub.3, COOR, and CONR.sub.2, where R is H or alkyl,
cycloalkyl, alkenyl, alkynyl, aryl, or heteroaryl. Where the
substituted atom is C, the substituents may include, in addition to
the substituents listed above, halo, OOCR, NROCR, where R is H or a
substituent set forth above.
"Alkyl" refers to a straight or branched hydrocarbon chain group
consisting solely of carbon and hydrogen atoms, containing no
unsaturation and including, for example, from one to ten carbon
atoms, and which is attached to the rest of the molecule by a
single bond. Unless stated otherwise specifically in the
specification, the alkyl group may be optionally substituted by one
or more substituents as described herein. Unless stated otherwise
specifically herein, it is understood that the substitution can
occur on any carbon of the alkyl group.
"Alkenyl" refers to a straight or branched hydrocarbon chain group
consisting solely of carbon and hydrogen atoms, containing at least
one double bond and including, for example, from two to ten carbon
atoms, and which is attached to the rest of the molecule by a
single bond or a double bond. Unless stated otherwise specifically
in the specification, the alkenyl group may be optionally
substituted by one or more substituents as described herein. Unless
stated otherwise specifically herein, it is understood that the
substitution can occur on any carbon of the alkenyl group.
"Alkynyl" refers to a straight or branched hydrocarbon chain group
consisting solely of carbon and hydrogen atoms, containing at least
one triple bond and including, for example, from two to ten carbon
atoms. Unless stated otherwise specifically in the specification,
the alkenyl group may be optionally substituted by one or more
substituents as described herein.
"Aryl" refers to a phenyl or naphthyl group, including for example,
5-12 members. Unless stated otherwise specifically herein, the term
"aryl" is meant to include aryl groups optionally substituted by
one or more substituents as described herein.
"Arylalkyl" refers to a group of the formula --R.sub.aR.sub.b where
R.sub.a is an alkyl group as described herein and R.sub.b is one or
more aryl moieties as described herein. The aryl group(s) may be
optionally substituted as described herein.
"Arylalkenyl" refers to a group of the formula --R.sub.cR.sub.b
where R.sub.a is an alkenyl moiety as described herein and R.sub.b
is one or more aryl groups as described herein. The aryl group(s)
and the alkenyl group may be optionally substituted as described
herein.
"Acyl" refers to a group of the formula --C(O)R.sub.a, where
R.sub.a is an alkyl group as described herein. The alkyl group(s)
may be optionally substituted as described herein.
"Arylacyl" refers to a group of the formula --C(O)R.sub.b, where
R.sub.b is an aryl group as described herein. The aryl group(s) may
be optionally substituted as described herein.
"Cycloalkyl" refers to a stable monovalent monocyclic, bicyclic or
tricyclic hydrocarbon group consisting solely of carbon and
hydrogen atoms, having for example from 3 to 15 carbon atoms, and
which is saturated and attached to the rest of the molecule by a
single bond. Unless otherwise stated specifically herein, the term
"cycloalkyl" is meant to include cycloalkyl groups which are
optionally substituted as described herein.
By a "ring structure" is meant a cycloalkyl, aryl, heteroaryl, or
any cyclic structure that may be optionally substituted.
"Optional" or "optionally" means that the subsequently described
event of circumstances may or may not occur, and that the
description includes instances where said event or circumstance
occurs and instances in which it does not. For example, "optionally
substituted alkyl" means that the alkyl group may or may not be
substituted and that the description includes both substituted
alkyl groups and alkyl groups having no substitution. Examples of
optionally substituted alkyl groups include, without limitation,
methyl, ethyl, propyl, etc. and including cycloalkyls such as
cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl,
etc.; examples of optionally substituted alkenyl groups include
allyl, crotyl, 2-pentenyl, 3-hexenyl, 2-cyclopentenyl,
2-cyclohexenyl, 2-cyclopentenylmethyl, 2-cyclohexenylmethyl, etc.
In some embodiments, optionally substituted alkyl and alkenyl
groups include C.sub.1-6 alkyls or alkenyls.
"Halo" refers to bromo, chloro, fluoro, iodo, etc. In some
embodiments, suitable halogens include fluorine or chlorine.
An amino group may also be substituted once or twice (to form a
secondary or tertiary amine) with a group such as an optionally
substituted alkyl group including C.sub.1-10alkyl (e.g., methyl,
ethyl propyl etc.); an optionally substituted alkenyl group such as
allyl, crotyl, 2-pentenyl, 3-hexenyl, etc., or an optionally
substituted cycloalkyl group such as cyclopropyl, cyclobutyl,
cyclopentyl, cyclohexyl, cycloheptyl, etc. In these cases,
C.sub.1-6 alkyl, alkenyl and cycloalkyl are preferred. The amine
group may also be optionally substituted with an aromatic or
heterocyclic group, aralkyl (e.g., phenylC.sub.1-4alkyl) or
heteroalkyl for example, phenyl, pyridine, phenylmethyl (benzyl),
phenethyl, pyridinylmethyl, pyridinylethyl, etc. The heterocyclic
group may be a 5 or 6 membered ring containing 1-4 heteroatoms.
An amino group may be substituted with an optionally substituted
C.sub.2-4 alkanoyl, e.g., acetyl, propionyl, butyryl, isobutyryl
etc., or a C.sub.1-4alkylsulfonyl (e.g., methanesulfonyl,
ethanesulfonyl, etc.) or a carbonyl or sulfonyl substituted
aromatic or heterocyclic ring, e.g., benzenesulfonyl, benzoyl,
pyridinesulfonyl, pyridinecarbonyl etc. The heterocycles are as
described herein.
Examples of optionally substituted carbonyl groups, or sulfonyl
groups include optionally substituted forms of such groups formed
from various hydrocarbyls such as alkyl, alkenyl and 5- to
6-membered monocyclic aromatic group (e.g., phenyl, pyridyl, etc.),
as described herein.
Therapeutic Indications
The invention provides methods of treating conditions that are
modulated, directly or indirectly, by an O-GlcNAcase enzyme or by
O-GlcNAc-modified protein levels, for example, a condition that is
benefited by inhibition of an O-GlcNAcase enzyme or by an elevation
of O-GlcNAc-modified protein levels. Such conditions include,
without limitation, Glaucoma, Schizophrenia, tauopathies, such as
Alzheimer's disease, neurodegenerative diseases, cardiovascular
diseases, diseases associated with inflammation, diseases
associated with immunosuppression and cancers. The compounds of the
invention are also useful in the treatment of diseases or disorders
related to deficiency or over-expression of O-GlcNAcase or
accumulation or depletion of O-GlcNAc, or any disease or disorder
responsive to glycosidase inhibition therapy. Such diseases and
disorders include, but are not limited to, Glaucoma, Schizophrenia,
neurodegenerative disorders, such as Alzheimer's disease (AD), or
cancer. Such diseases and disorders may also include diseases or
disorders related to the accumulation or deficiency in the enzyme
OGT. Also included is a method of protecting or treating target
cells expressing proteins that are modified by O-GlcNAc residues,
the dysregulation of which modification results in disease or
pathology. The term "treating" as used herein includes treatment,
prevention, and amelioration.
In alternative embodiments, the invention provides methods of
enhancing or elevating levels of protein O-GlcNAc modification in
animal subjects, such as, veterinary and human subjects. This
elevation of O-GlcNAc levels can be useful for the prevention or
treatment of Alzheimer's disease; prevention or treatment of other
neurodegenerative diseases (e.g. Parkinson's disease, Huntington's
disease); providing neuroprotective effects; preventing damage to
cardiac tissue; and treating diseases associated with inflammation
or immunosuppression.
In alternative embodiments, the invention provides methods of
selectively inhibiting an O-GlcNAcase enzyme in animal subjects,
such as veterinary and human subjects.
In alternative embodiments, the invention provides methods of
inhibiting phosphorylation of tau polypeptides, or inhibiting
formation of NFTs, in animal subjects, such as, veterinary and
human subjects. Accordingly, the compounds of the invention may be
used to study and treat AD and other tauopathies.
In general, the methods of the invention are effected by
administering a compound according to the invention to a subject in
need thereof, or by contacting a cell or a sample with a compound
according to the invention, for example, a pharmaceutical
composition comprising a therapeutically effective amount of the
compound according to Formula (I). More particularly, they are
useful in the treatment of a disorder in which the regulation of
O-GlcNAc protein modification is implicated, or any condition as
described herein. Disease states of interest include Alzheimer's
disease (AD) and related neurodegenerative tauopathies, in which
abnormal hyperphosphorylation of the microtubule-associated protein
tau is involved in disease pathogenesis. In some embodiments, the
compounds may be used to block hyperphosphorylation of tau by
maintaining elevated levels of O-GlcNAc on tau, thereby providing
therapeutic benefit.
Tauopathies that may be treated with the compounds of the invention
include: Alzheimer's disease, Amyotrophic lateral sclerosis (ALS),
Amyotrophic lateral sclerosis with cognitive impairment (ALSci),
Argyrophilic grain dementia, Bluit disease, Corticobasal
degeneration (CBD), Dementia pugilistica, Diffuse neurofibrillary
tangles with calcification, Down's syndrome, Familial British
dementia, Familial Danish dementia, Frontotemporal dementia with
parkinsonism linked to chromosome 17 (FTDP-17),
Gerstmann-Straussler-Scheinker disease, Guadeloupean parkinsonism,
Hallevorden-Spatz disease (neurodegeneration with brain iron
accumulation type 1), Multiple system atrophy, Myotonic dystrophy,
Niemann-Pick disease (type C), Pallido-ponto-nigral degeneration,
Parkinsonism-dementia complex of Guam, Pick's disease (PiD),
Post-encephalitic parkinsonism (PEP), Prion diseases (including
Creutzfeldt-Jakob Disease (CJD), Variant Creutzfeldt-Jakob Disease
(vCJD), Fatal Familial Insomnia, and Kuru), Progressive
supercortical gliosis, Progressive supranuclear palsy (PSP),
Richardson's syndrome, Subacute sclerosing panencephalitis, and
Tangle-only dementia.
The compounds of this invention are also useful in the treatment of
conditions associate with tissue damage or stress, stimulating
cells, or promoting differentiation of cells. Accordingly, in some
embodiments, the compounds of this invention may be used to provide
therapeutic benefit in a variety of conditions or medical
procedures involving stress in cardiac tissue, including but not
limited to: ischemia; hemorrhage; hypovolemic shock; myocardial
infarction; an interventional cardiology procedure; cardiac bypass
surgery; fibrinolytic therapy; angioplasty; and stent
placement.
Compounds that selectively inhibit O-GlcNAcase activity may be used
for the treatment of diseases that are associated with
inflammation, including but not limited to, inflammatory or
allergic diseases such as asthma, allergic rhinitis,
hypersensitivity lung diseases, hypersensitivity pneumonitis,
eosinophilic pneumonias, delayed-type hypersensitivity,
atherosclerosis, interstitial lung disease (ILD) (e.g., idiopathic
pulmonary fibrosis, or ILD associated with rheumatoid arthritis,
systemic lupus erythematosus, ankylosing spondylitis, systemic
sclerosis, Sjogren's syndrome, polymyositis or dermatomyositis);
systemic anaphylaxis or hypersensitivity responses, drug allergies,
insect sting allergies; autoimmune diseases, such as rheumatoid
arthritis, psoriatic arthritis, multiple sclerosis, systemic lupus
erythematosus, myastenia gravis, glomerulonephritis, autoimmune
thyroiditis, graft rejection, including allograft rejection or
graft-versus-host disease; inflammatory bowel diseases, such as
Crohn's disease and ulcerative colitis; spondyloarthropathies;
scleroderma; psoriasis (including T-cell mediated psoriasis) and
inflammatory dermatoses such as dermatitis, eczema, atopic
dermatitis, allergic contact dermatitis, urticaria; vasculitis
(e.g., necrotizing, cutaneous, and hypersensitivity vasculitis);
eosinphilic myotis, eosiniphilic fasciitis; and cancers.
In addition, compounds that affects levels of protein O-GlcNAc
modification may be used for the treatment of diseases associated
with immunosuppression, such as in individuals undergoing
chemotherapy, radiation therapy, enhanced wound healing and burn
treatment, therapy for autoimmune disease or other drug therapy
(e.g., corticosteroid therapy) or combination of conventional drugs
used in the treatment of autoimmune diseases and
graft/transplantation rejection, which causes immunosuppression; or
immunosuppression due to congenital deficiency in receptor function
or other causes.
The compounds of the invention may be useful for treatment of
neurodegenerative diseases, including Parkinson's disease and
Huntington's disease. Other conditions that may be treated are
those triggered, affected, or in any other way correlated with
levels of O-GlcNAc post-translational protein modification. It is
expected that the compounds of this invention may be useful for the
treatment of such conditions and in particular, but not limited to,
the following for which a association with O-GlcNAc levels on
proteins has been established: graft rejection, in particular but
not limited to solid organ transplants, such as heart, lung, liver,
kidney, and pancreas transplants (e.g. kidney and lung allografts);
cancer, in particular but not limited to cancer of the breast,
lung, prostate, pancreas, colon, rectum, bladder, kidney, ovary; as
well as non-Hodgkin's lymphoma and melanoma; epilepsy, pain, or
stroke, e.g., for neuroprotection following a stroke.
Pharmaceutical & Veterinary Compositions, Dosages, and
Administration
Pharmaceutical compositions including compounds according to the
invention, or for use according to the invention, are contemplated
as being within the scope of the invention. In some embodiments,
pharmaceutical compositions including an effective amount of a
compound of Formula (I) are provided.
The compounds of formula (I) and their pharmaceutically acceptable
salts, stereoisomers, solvates, and derivatives are useful because
they have pharmacological activity in animals, including humans. In
some embodiments, the compounds according to the invention are
stable in plasma, when administered to a subject.
In some embodiments, compounds according to the invention, or for
use according to the invention, may be provided in combination with
any other active agents or pharmaceutical compositions where such
combined therapy is useful to modulate O-GlcNAcase activity, for
example, to treat neurodegenerative, inflammatory, cardiovascular,
or immunoregulatory diseases, or any condition described herein. In
some embodiments, compounds according to the invention, or for use
according to the invention, may be provided in combination with one
or more agents useful in the prevention or treatment of Alzheimer's
disease. Examples of such agents include, without limitation,
acetylcholine esterase inhibitors (AChEIs) such as Aricept.RTM.
(Donepezil), Exelon.RTM. (Rivastigmine), Razadyne.RTM. (Razadyne
ER.RTM., Reminyl.RTM., Nivalin.RTM., Galantamine), Cognex.RTM.
(Tacrine), Dimebon, Huperzine A, Phenserine, Debio-9902 SR (ZT-1
SR), Zanapezil (TAK0147), ganstigmine, NP7557, etc.; NMDA receptor
antagonists such as Namenda.RTM. (Axura.RTM., Akatinol.RTM.,
Ebixa.RTM., Memantine), Dimebon, SGS-742, Neramexane, Debio-9902 SR
(ZT-1 SR), etc.; gamma-secretase inhibitors and/or modulators such
as Flurizan.TM. (Tarenflurbil, MPC-7869, R-flurbiprofen), LY450139,
MK 0752, E2101, BMS-289948, BMS-299897, BMS-433796, LY-411575,
GSI-136, etc.; beta-secretase inhibitors such as ATG-Z1, CTS-21166,
etc.; alpha-secretase activators, such as NGX267, etc;
amyloid-.beta. aggregation and/or fibrillization inhibitors such as
Alzhemed.TM. (3APS, Tramiprosate, 3-amino-1-propanesulfonic acid),
AL-108, AL-208, AZD-103, PBT2, Cereact, ONO-2506PO, PPI-558, etc.;
tau aggregation inhibitors such as methylene blue, etc.;
microtubule stabilizers such as AL-108, AL-208, paclitaxel, etc.;
RAGE inhibitors, such as TTP488, etc.; 5-HT1a receptor antagonists,
such as Xaliproden, Lecozotan, etc.; 5-HT4 receptor antagonists,
such as PRX-03410, etc.; kinase inhibitors such as SRN-003-556,
amfurindamide, LiCl, AZD1080, NP031112, SAR-502250, etc. humanized
monoclonal anti-.beta. antibodies such as Bapineuzumab (AAB-001),
LY2062430, RN1219, ACU-5A5, etc.; amyloid vaccines such as AN-1792,
ACC-001 neuroprotective agents such as Cerebrolysin, AL-108,
AL-208, Huperzine A, etc.; L-type calcium channel antagonists such
as MEM-1003, etc.; nicotinic receptor antagonists, such as AZD3480,
GTS-21, etc.; nicotinic receptor agonists, such as MEM 3454,
Nefiracetam, etc.; peroxisome proliferator-activated receptor
(PPAR) gamma agonists such as Avandia.RTM. (Rosglitazone), etc.;
phosphodiesterase IV (PDE4) inhibitors, such as MK-0952, etc.;
hormone replacement therapy such as estrogen (Premarin), etc.;
monoamine oxidase (MAO) inhibitors such as NS2330, Rasagiline
(Azilect.RTM.), TVP-1012, etc.; AMPA receptor modulators such as
Ampalex (CX 516), etc.; nerve growth factors or NGF potentiators,
such as CERE-110 (AAV-NGF), T-588, T-817MA, etc.; agents that
prevent the release of luteinizing hormone (LH) by the pituitary
gland, such as leuoprolide (VP-4896), etc.; GABA receptor
modulators such as AC-3933, NGD 97-1, CP-457920, etc.;
benzodiazepine receptor inverse agonists such as SB-737552
(S-8510), AC-3933, etc.; noradrenaline-releasing agents such as
T-588, T-817MA, etc.
It is to be understood that combination of compounds according to
the invention, or for use according to the invention, with
Alzheimer's agents is not limited to the examples described herein,
but includes combination with any agent useful for the treatment of
Alzheimer's disease. Combination of compounds according to the
invention, or for use according to the invention, and other
Alzheimer's agents may be administered separately or in
conjunction. The administration of one agent may be prior to,
concurrent to, or subsequent to the administration of other
agent(s).
In alternative embodiments, the compounds may be supplied as
"prodrugs" or protected forms, which release the compound after
administration to a subject. For example, the compound may carry a
protective group which is split off by hydrolysis in body fluids,
e.g., in the bloodstream, thus releasing the active compound or is
oxidized or reduced in body fluids to release the compound.
Accordingly, a "prodrug" is meant to indicate a compound that may
be converted under physiological conditions or by solvolysis to a
biologically active compound of the invention. Thus, the term
"prodrug" refers to a metabolic precursor of a compound of the
invention that is pharmaceutically acceptable. A prodrug may be
inactive when administered to a subject in need thereof, but is
converted in vivo to an active compound of the invention. Prodrugs
are typically rapidly transformed in vivo to yield the parent
compound of the invention, for example, by hydrolysis in blood. The
prodrug compound often offers advantages of solubility, tissue
compatibility or delayed release in a subject.
The term "prodrug" is also meant to include any covalently bonded
carriers which release the active compound of the invention in vivo
when such prodrug is administered to a subject. Prodrugs of a
compound of the invention may be prepared by modifying functional
groups present in the compound of the invention in such a way that
the modifications are cleaved, either in routine manipulation or in
vivo, to the parent compound of the invention. Prodrugs include
compounds of the invention wherein a hydroxy, amino or mercapto
group is bonded to any group that, when the prodrug of the compound
of the invention is administered to a mammalian subject, cleaves to
form a free hydroxy, free amino or free mercapto group,
respectively. Examples of prodrugs include, but are not limited to,
acetate, formate and benzoate derivatives of alcohol and acetamide,
formamide, and benzamide derivatives of amine functional groups in
the compounds of the invention and the like.
Additional examples of prodrugs for the compounds of the invention
include acetonide derivatives (also known as isopropylidine
derivatives) in which two OR.sup.1 groups in Formula (I) may be
linked in a ring, for example, as in Formulae (II) and (III) shown
below. Such acetonide groups may be cleaved in vivo to liberate the
parent compound of the invention, making these acetonide
derivatives prodrugs.
##STR00030##
A discussion of prodrugs may be found in "Smith and Williams'
Introduction to the Principles of Drug Design," H. J. Smith,
Wright, Second Edition, London (1988); Bundgard, H., Design of
Prodrugs (1985), pp. 7-9, 21-24 (Elsevier, Amsterdam); The Practice
of Medicinal Chemistry, Camille G. Wermuth et al., Ch 31, (Academic
Press, 1996); A Textbook of Drug Design and Development, P.
Krogsgaard-Larson and H. Bundgaard, eds. Ch 5, pgs 113 191 (Harwood
Academic Publishers, 1991); Higuchi, T., et al., "Pro-drugs as
Novel Delivery Systems," A.C.S. Symposium Series, Vol. 14; or in
Bioreversible Carriers in Drug Design, ed. Edward B. Roche,
American Pharmaceutical Association and Pergamon Press, 1987, all
of which are incorporated in full by reference herein.
Suitable prodrug forms of the compounds of the invention include
embodiments in which R.sup.1 is C(O)R, where R is optionally
substituted alkyl, alkenyl, alkynyl, aryl, or heteroaryl. In these
cases the ester groups may be hydrolyzed in vivo (e.g. in bodily
fluids), releasing the active compounds in which R.sup.1 is H.
Preferred prodrug embodiments of the invention are the compounds of
Formula (I) where R.sup.1 is C(O)CH.sub.3.
Compounds according to the invention, or for use according to the
invention, can be provided alone or in combination with other
compounds in the presence of a liposome, an adjuvant, or any
pharmaceutically acceptable carrier, diluent or excipient, in a
form suitable for administration to a subject such as a mammal, for
example, humans, cattle, sheep, etc. If desired, treatment with a
compound according to the invention may be combined with more
traditional and existing therapies for the therapeutic indications
described herein. Compounds according to the invention may be
provided chronically or intermittently. "Chronic" administration
refers to administration of the compound(s) in a continuous mode as
opposed to an acute mode, so as to maintain the initial therapeutic
effect (activity) for an extended period of time. "Intermittent"
administration is treatment that is not consecutively done without
interruption, but rather is cyclic in nature. The terms
"administration," "administrable," or "administering" as used
herein should be understood to mean providing a compound of the
invention to the subject in need of treatment.
"Pharmaceutically acceptable carrier, diluent or excipient"
includes without limitation any adjuvant, carrier, excipient,
glidant, sweetening agent, diluent, preservative, dye/colorant,
flavor enhancer, surfactant, wetting agent, dispersing agent,
suspending agent, stabilizer, isotonic agent, solvent, or
emulsifier that has been approved, for example, by the United
States Food and Drug Administration or other governmental agency as
being acceptable for use in humans or domestic animals.
The compounds of the present invention may be administered in the
form of pharmaceutically acceptable salts. In such cases,
pharmaceutical compositions in accordance with this invention may
comprise a salt of such a compound, preferably a physiologically
acceptable salt, which are known in the art. In some embodiments,
the term "pharmaceutically acceptable salt" as used herein means an
active ingredient comprising compounds of Formula 1 used in the
form of a salt thereof, particularly where the salt form confers on
the active ingredient improved pharmacokinetic properties as
compared to the free form of the active ingredient or other
previously disclosed salt form.
A "pharmaceutically acceptable salt" includes both acid and base
addition salts. A "pharmaceutically acceptable acid addition salt"
refers to those salts which retain the biological effectiveness and
properties of the free bases, which are not biologically or
otherwise undesirable, and which are formed with inorganic acids
such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric
acid, phosphoric acid and the like, and organic acids such as
acetic acid, trifluoroacetic acid, propionic acid, glycolic acid,
pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic
acid, fumaric acid, tartaric acid, citric acid, benzoic acid,
cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic
acid, p-toluenesulfonic acid, salicylic acid, and the like.
A "pharmaceutically acceptable base addition salt" refers to those
salts which retain the biological effectiveness and properties of
the free acids, which are not biologically or otherwise
undesirable. These salts are prepared from addition of an inorganic
base or an organic base to the free acid. Salts derived from
inorganic bases include, but are not limited to, the sodium,
potassium, lithium, ammonium, calcium, magnesium, iron, zinc,
copper, manganese, aluminum salts and the like. Preferred inorganic
salts are the ammonium, sodium, potassium, calcium, and magnesium
salts. Salts derived from organic bases include, but are not
limited to, salts of primary, secondary, and tertiary amines,
substituted amines including naturally occurring substituted
amines, cyclic amines and basic ion exchange resins, such as
isopropylamine, trimethylamine, diethylamine, triethylamine,
tripropylamine, ethanolamine, 2-dimethylaminoethanol,
2-diethylaminoethanol, dicyclohexylamine, lysine, arginine,
histidine, caffeine, procaine, hydrab amine, choline, betaine,
ethylenediamine, glucosamine, methylglucamine, theobromine,
purines, piperazine, piperidine, N-ethylpiperidine, polyamine
resins and the like. Particularly preferred organic bases are
isopropylamine, diethylamine, ethanolamine, trimethylamine,
dicyclohexylamine, choline and caffeine.
Thus, the term "pharmaceutically acceptable salt" encompasses all
acceptable salts including but not limited to acetate,
lactobionate, benzenesulfonate, laurate, benzoate, malate,
bicarbonate, maleate, bisulfate, mandelate, bitartarate, mesylate,
borate, methylbromide, bromide, methylnitrite, calcium edetate,
methylsulfate, camsylate, mucate, carbonate, napsylate, chloride,
nitrate, clavulanate, N-methylglucamine, citrate, ammonium salt,
dihydrochloride, oleate, edetate, oxalate, edisylate, pamoate
(embonate), estolate, palmitate, esylate, pantothenate, fumarate,
phosphate/diphosphate, gluceptate, polygalacturonate, gluconate,
salicylate, glutame, stearate, glycollylarsanilate, sulfate,
hexylresorcinate, subacetate, hydradamine, succinate, hydrobromide,
tannate, hydrochloride, tartrate, hydroxynaphthoate, teoclate,
iodide, tosylate, isothionate, triethiodide, lactate, panoate,
valerate, and the like.
Pharmaceutically acceptable salts of the compounds of the present
invention can be used as a dosage for modifying solubility or
hydrolysis characteristics, or can be used in sustained release or
prodrug formulations. Also, pharmaceutically acceptable salts of
the compounds of this invention may include those formed from
cations such as sodium, potassium, aluminum, calcium, lithium,
magnesium, zinc, and from bases such as ammonia, ethylenediamine,
N-methyl-glutamine, lysine, arginine, ornithine, choline,
N,N'-dibenzylethylene-diamine, chloroprocaine, diethanolamine,
procaine, N-benzylphenethyl-amine, diethylamine, piperazine,
tris(hydroxymethyl)aminomethane, and tetramethylammonium
hydroxide.
Pharmaceutical formulations will typically include one or more
carriers acceptable for the mode of administration of the
preparation, be it by injection, inhalation, topical
administration, lavage, or other modes suitable for the selected
treatment. Suitable carriers are those known in the art for use in
such modes of administration.
Suitable pharmaceutical compositions may be formulated by means
known in the art and their mode of administration and dose
determined by the skilled practitioner. For parenteral
administration, a compound may be dissolved in sterile water or
saline or a pharmaceutically acceptable vehicle used for
administration of non-water soluble compounds such as those used
for vitamin K. For enteral administration, the compound may be
administered in a tablet, capsule or dissolved in liquid form. The
table or capsule may be enteric coated, or in a formulation for
sustained release. Many suitable formulations are known, including,
polymeric or protein microparticles encapsulating a compound to be
released, ointments, gels, hydrogels, or solutions which can be
used topically or locally to administer a compound. A sustained
release patch or implant may be employed to provide release over a
prolonged period of time. Many techniques known to skilled
practitioners are described in Remington: the Science &
Practice of Pharmacy by Alfonso Gennaro, 20.sup.th ed., Williams
& Wilkins, (2000). Formulations for parenteral administration
may, for example, contain excipients, polyalkylene glycols such as
polyethylene glycol, oils of vegetable origin, or hydrogenated
naphthalenes. Biocompatible, biodegradable lactide polymer,
lactide/glycolide copolymer, or polyoxyethylene-polyoxypropylene
copolymers may be used to control the release of the compounds.
Other potentially useful parenteral delivery systems for modulatory
compounds include ethylene-vinyl acetate copolymer particles,
osmotic pumps, implantable infusion systems, and liposomes.
Formulations for inhalation may contain excipients, for example,
lactose, or may be aqueous solutions containing, for example,
polyoxyethylene-9-lauryl ether, glycocholate and deoxycholate, or
may be oily solutions for administration in the form of nasal
drops, or as a gel.
The compounds or pharmaceutical compositions according to the
present invention may be administered by oral or non-oral, e.g.,
intramuscular, intraperitoneal, intravenous, intracisternal
injection or infusion, subcutaneous injection, transdermal or
transmucosal routes. In some embodiments, compounds or
pharmaceutical compositions in accordance with this invention or
for use in this invention may be administered by means of a medical
device or appliance such as an implant, graft, prosthesis, stent,
etc. Implants may be devised which are intended to contain and
release such compounds or compositions. An example would be an
implant made of a polymeric material adapted to release the
compound over a period of time. The compounds may be administered
alone or as a mixture with a pharmaceutically acceptable carrier
e.g., as solid formulations such as tablets, capsules, granules,
powders, etc.; liquid formulations such as syrups, injections,
etc.; injections, drops, suppositories, pessaryies. In some
embodiments, compounds or pharmaceutical compositions in accordance
with this invention or for use in this invention may be
administered by inhalation spray, nasal, vaginal, rectal,
sublingual, or topical routes and may be formulated, alone or
together, in suitable dosage unit formulations containing
conventional non-toxic pharmaceutically acceptable carriers,
adjuvants and vehicles appropriate for each route of
administration.
The compounds of the invention may be used to treat animals,
including mice, rats, horses, cattle, sheep, dogs, cats, and
monkeys. However, compounds of the invention can also be used in
other organisms, such as avian species (e.g., chickens). The
compounds of the invention may also be effective for use in humans.
The term "subject" or alternatively referred to herein as "patient"
is intended to be referred to an animal, preferably a mammal, most
preferably a human, who has been the object of treatment,
observation or experiment. However, the compounds, methods and
pharmaceutical compositions of the present invention may be used in
the treatment of animals. Accordingly, as used herein, a "subject"
may be a human, non-human primate, rat, mouse, cow, horse, pig,
sheep, goat, dog, cat, etc. The subject may be suspected of having
or at risk for having a condition requiring modulation of
O-GlcNAcase activity.
An "effective amount" of a compound according to the invention
includes a therapeutically effective amount or a prophylactically
effective amount. A "therapeutically effective amount" refers to an
amount effective, at dosages and for periods of time necessary, to
achieve the desired therapeutic result, such as inhibition of an
O-GlcNAcase, elevation of O-GlcNAc levels, inhibition of tau
phosphorylation, or any condition described herein. A
therapeutically effective amount of a compound may vary according
to factors such as the disease state, age, sex, and weight of the
individual, and the ability of the compound to elicit a desired
response in the individual. Dosage regimens may be adjusted to
provide the optimum therapeutic response. A therapeutically
effective amount is also one in which any toxic or detrimental
effects of the compound are outweighed by the therapeutically
beneficial effects. A "prophylactically effective amount" refers to
an amount effective, at dosages and for periods of time necessary,
to achieve the desired prophylactic result, such as inhibition of
an O-GlcNAcase, elevation of O-GlcNAc levels, inhibition of tau
phosphorylation, or any condition described herein. Typically, a
prophylactic dose is used in subjects prior to or at an earlier
stage of disease, so that a prophylactically effective amount may
be less than a therapeutically effective amount. A suitable range
for therapeutically or prophylactically effective amounts of a
compound may be any integer from 0.1 nM-0.1M, 0.1 nM-0.05M, 0.05
nM-15 .mu.M or 0.01 nM-10 .mu.M.
In alternative embodiments, in the treatment or prevention of
conditions which require modulation of O-GlcNAcase activity, an
appropriate dosage level will generally be about 0.01 to 500 mg per
kg subject body weight per day, and can be administered in singe or
multiple doses. In some embodiments, the dosage level will be about
0.1 to about 250 mg/kg per day. It will be understood that the
specific dose level and frequency of dosage for any particular
patient may be varied and will depend upon a variety of factors
including the activity of the specific compound used, the metabolic
stability and length of action of that compound, the age, body
weight, general health, sex, diet, mode and time of administration,
rate of excretion, drug combination, the severity of the particular
condition, and the patient undergoing therapy.
It is to be noted that dosage values may vary with the severity of
the condition to be alleviated. For any particular subject,
specific dosage regimens may be adjusted over time according to the
individual need and the professional judgement of the person
administering or supervising the administration of the
compositions. Dosage ranges set forth herein are exemplary only and
do not limit the dosage ranges that may be selected by medical
practitioners. The amount of active compound(s) in the composition
may vary according to factors such as the disease state, age, sex,
and weight of the subject. Dosage regimens may be adjusted to
provide the optimum therapeutic response. For example, a single
bolus may be administered, several divided doses may be
administered over time or the dose may be proportionally reduced or
increased as indicated by the exigencies of the therapeutic
situation. It may be advantageous to formulate parenteral
compositions in dosage unit form for ease of administration and
uniformity of dosage. In general, compounds of the invention should
be used without causing substantial toxicity, and as described
herein, the compounds exhibit a suitable safety profile for
therapeutic use. Toxicity of the compounds of the invention can be
determined using standard techniques, for example, by testing in
cell cultures or experimental animals and determining the
therapeutic index, i.e., the ratio between the LD50 (the dose
lethal to 50% of the population) and the LD100 (the dose lethal to
100% of the population). In some circumstances however, such as in
severe disease conditions, it may be necessary to administer
substantial excesses of the compositions.
Other Uses and Assays
A compound of Formula (I) may be used in screening assays for
compounds which modulate the activity of glycosidase enzymes,
preferably the O-GlcNAcase enzyme. The ability of a test compound
to inhibit O-GlcNAcase-dependent cleavage of O-GlcNAc from a model
substrate may be measured using any assays, as described herein or
known to one of ordinary skill in the art. For example, a
fluoresence or UV-based assay known in the art may be used. A "test
compound" is any naturally-occurring or artificially-derived
chemical compound. Test compounds may include, without limitation,
peptides, polypeptides, synthesised organic molecules, naturally
occurring organic molecules, and nucleic acid molecules. A test
compound can "compete" with a known compound such as a compound of
Formula (I) by, for example, interfering with inhibition of
O-GlcNAcase-dependent cleavage of O-GlcNAc or by interfering with
any biological response induced by a compound of Formula (I).
Generally, a test compound can exhibit any value between 10% and
200%, or over 500%, modulation when compared to a compound of
Formula (I) or other reference compound. For example, a test
compound may exhibit at least any positive or negative integer from
10% to 200% modulation, or at least any positive or negative
integer from 30% to 150% modulation, or at least any positive or
negative integer from 60% to 100% modulation, or any positive or
negative integer over 100% modulation. A compound that is a
negative modulator will in general decrease modulation relative to
a known compound, while a compound that is a positive modulator
will in general increase modulation relative to a known
compound.
In general, test compounds are identified from large libraries of
both natural products or synthetic (or semi-synthetic) extracts or
chemical libraries according to methods known in the art. Those
skilled in the field of drug discovery and development will
understand that the precise source of test extracts or compounds is
not critical to the method(s) of the invention. Accordingly,
virtually any number of chemical extracts or compounds can be
screened using the exemplary methods described herein. Examples of
such extracts or compounds include, but are not limited to, plant-,
fungal-, prokaryotic- or animal-based extracts, fermentation
broths, and synthetic compounds, as well as modification of
existing compounds. Numerous methods are also available for
generating random or directed synthesis (e.g., semi-synthesis or
total synthesis) of any number of chemical compounds, including,
but not limited to, saccharide-, lipid-, peptide-, and nucleic
acid-based compounds. Synthetic compound libraries are commercially
available. Alternatively, libraries of natural compounds in the
form of bacterial, fungal, plant, and animal extracts are
commercially available from a number of sources, including Biotics
(Sussex, UK), Xenova (Slough, UK), Harbor Branch Oceanographic
Institute (Ft. Pierce, Fla., USA), and PharmaMar, Mass., USA. In
addition, natural and synthetically produced libraries are
produced, if desired, according to methods known in the art, e.g.,
by standard extraction and fractionation methods. Furthermore, if
desired, any library or compound is readily modified using standard
chemical, physical, or biochemical methods.
When a crude extract is found to modulate inhibition of
O-GlcNAcase-dependent cleavage of O-GlcNAc, or any biological
response induced by a compound of Formula (I), further
fractionation of the positive lead extract is necessary to isolate
chemical constituents responsible for the observed effect. Thus,
the goal of the extraction, fractionation, and purification process
is the careful characterization and identification of a chemical
entity within the crude extract having O-GlcNAcase-inhibitory
activities. The same assays described herein for the detection of
activities in mixtures of compounds can be used to purify the
active component and to test derivatives thereof. Methods of
fractionation and purification of such heterogeneous extracts are
known in the art. If desired, compounds shown to be useful agents
for treatment are chemically modified according to methods known in
the art. Compounds identified as being of therapeutic,
prophylactic, diagnostic, or other value may be subsequently
analyzed using a suitable animal model, as described herein on
known in the art.
In some embodiments, the compounds described herein (e.g., the
compounds of Formula I) or test compounds may be analyzed using
established cellular.sup.118-120 and/or transgenic animal models of
disease.sup.32,33 and the ability of the compounds to, for example,
block the formation of toxic tau species determined. Such analyses
may be used for example to determine or confirm the efficacy of the
compounds in treating or preventing pathology associated with the
accumulation of toxic tau species (for example, Alzheimer's disease
and other tauopathies).
In some embodiments, the compounds described herein (e.g., the
compounds of Formula I) or test compounds may be analyzed using
established cellular stress assays.sup.105,116,117 and/or animal
models of ischemia-reperfusion.sup.70,114 or
trauma-hemorrhage..sup.72,112,115 Such analyses may be used for
example to determine or confirm the efficacy of the compounds in
treating or preventing pathology associated with cellular stress
(including ischemia, hemorrhage, hypovolemic shock, myocardial
infarction, and other cardiovascular disorders) or in treating or
preventing tissue damage or promoting functional recovery.
In some embodiments, the compounds are useful in the development of
animal models for studying diseases or disorders related to
deficiencies in O-GlcNAcase, over-expression of O-GlcNAcase,
accumulation of O-GlcNAc, depletion of O-GlcNAc, and for studying
treatment of diseases and disorders related to deficiency or
over-expression of O-GlcNAcase, or accumulation or depletion of
O-GlcNAc. Such diseases and disorders include neurodegenerative
diseases, including Alzheimer's disease, and cancer.
Various alternative embodiments and examples of the invention are
described herein. These embodiments and examples are illustrative
and should not be construed as limiting the scope of the
invention.
EXAMPLES
The following examples are intended to illustrate embodiments of
the invention and are not intended to be construed in a limiting
manner.
Example 1
Compounds 1 and 2 (allosamizoline) are prepared via known synthetic
methods..sup.121-131 For example, following the synthetic route of
Trost et al..sup.126 (Scheme 1), reaction of the diol A with 2
equiv. TsNCO followed by catalytic palladium provides the cyclic
carbamate B in 96% yield. Removal of the tosyl group with sodium in
napthalene gives C in 91% yield, which is then converted in
quantitative yield to the amino oxazoline D by reaction with
freshly-distilled methyl triflate followed by dimethyl amine.
Dihydroxylation of the double bond in D using trifluroperacetic
acid in TFA then warming with aqueous TFA, followed by benzyl
deprotection with hydrogenolysis provides allosamizoline 2 in 67%
yield. Numerous alternative synthetic routes are available for the
preparation of 2, as described in the references cited
herein..sup.121-131
##STR00031##
Example 2
Compounds of the invention having general structure F are prepared
according to the sequence described in Scheme 2. Thus, starting
from the intermediate C,.sup.126 treatment with methyl triflate
followed by the requisite primary or secondary amine (analagous to
the procedure described by Trost et al..sup.126) provides the amino
oxazolines E. Dihydroxylation of these materials followed by
deprotection using the same conditions described in Scheme 1
provides the desired products F.
##STR00032##
Example 3
Compounds of the invention having general structure K are prepared
according to the sequence described in Scheme 3. Thus, starting
from the intermediate G,.sup.132 treatment with the appropriate
isothiocyanate provides the ureas H. Mesylation of the alcohol
function in H provides intermediates I. Treatment of these
compounds with potassium iodide in acetone converts the mesylate to
an iodo function with inversion of stereochemistry; under the
reaction conditions, the thiourea cyclizes onto the iodo group to
give intermediates J. Deprotection of the TBDMS group in these
materials with TBAF, followed by benzyl deprotection using
hydrogenolysis furnishes the desired products K.
##STR00033##
Example 4
Compounds of the invention having general structure L are prepared
according to the sequence described in Scheme 4. Treatment of
compounds K with the appropriate secondary amine in water with
heating provided the desired products L (e.g., see Kinoshita et
al..sup.133)
##STR00034##
Example 5
Compound 2 (allosamizoline):
(3aR,4R,5R,6R,6aS)-2-(dimethylamino)-6-(hydroxymethyl)-4,5,6,6a-tetrahydr-
o-3aH-cyclopenta[d]oxazole-4,5-diol
##STR00035##
Compound 2 was purchased as the hydrochloride salt from a
commercial supplier. .sup.1H NMR (200 MHz, D.sub.2O) .delta.
2.25-2.40 (m, 1H), 3.00 (s, 3H), 3.02 (s, 3H), 3.59-3.87 (m, 3H),
4.02 (dd, 1H, J=7, 5 Hz), 4.27 (dd, 1H, J=9, 6 Hz), 5.26 (dd, 1H,
J=9, 5 Hz); .sup.13C NMR (50 MHz, D.sub.2O) .delta. 37.3, 37.5,
51.3, 59.3, 63.6, 74.8, 81.6, 86.7, 160.6; MS (EI, free base): m/z
217 (M+1).
Example 6
Assay for Determination of K.sub.1 Values for Inhibition of
O-GlcNAcase Activity
Experimental Procedure for Kinetic Analyses:
Enzymatic reactions are carried out in PBS buffer (pH 7.4) using
pNP-GlcNAc as a substrate (0.5 mM) and monitored continuously at
37.degree. C. at 400 nm using a Cary 3E UV-VIS spectrophotometer
equipped with a Peltier temperature controller. Reactions are
pre-heated in a 500 .mu.L quartz cuvette for approximately 5
minutes followed by addition of 10 .mu.L enzyme via syringe (final
enzyme concentration 0.002 mg/mL). Reaction velocities are
determined by linear regression of the linear region of the
reaction progress curve between the first and third minutes. An
inhibitor concentration range of 1/5 to 5 times K.sub.1 is used in
each case. When tested in this assay, many of the compounds
described herein exhibit K.sub.1 values for inhibition of
O-GlcNAcase in the range 1 nM-50 .mu.M. All K.sub.1 values are
determined using linear regression of Dixon plots.
Example 7
Assay for Determination of K.sub.1 Values for Inhibition of
.beta.-Hexosaminidase Activity
Experimental Procedure for Kinetic Analyses:
All enzymatic assays are carried out in triplicate at 37.degree. C.
using a stopped assay procedure by measuring the amount of
4-nitrophenolate liberated as determined by absorption measurements
at 400 nm. Reactions (50 .mu.L) are initiated by the addition, via
syringe, of enzyme (3 .mu.L). Time-dependent assay of
.beta.-hexosaminidase has revealed that the enzyme is stable in the
buffer over the period of the assay: 50 mM citrate, 100 mM NaCl,
0.1% BSA, pH 4.25. .beta.-hexosaminidase is used at a concentration
of 0.036 mg/mL with pNP-GlcNAc as a substrate at a concentration of
0.5 mM. The inhibitor is tested at five concentrations ranging from
5 times to 1/5 K.sub.1. K.sub.1 values are determined by linear
regression of data from Dixon plots.
When tested in this assay, many of the compounds described herein
exhibit K.sub.1 values for inhibition of .beta.-hexosaminidase in
the range 1 .mu.M-10 mM. For example, the K.sub.1 value for
inhibition of O-GlcNAcase shown in Table 3 was obtained for
compound 2. This K.sub.1 value was determined using linear
regression of a Dixon plot.
TABLE-US-00003 TABLE 3 Inhibition constant for O-GlcNAcase.
Compound O-GlcNAcase K.sub.I (.mu.M) 2 40
The selectivity ratio for inhibition of O-GlcNAcase over
.beta.-hexosaminidase is defined here as:
K.sub.1(.beta.-hexosaminidase)/K.sub.1(O-GlcNAcase) In general, the
compounds described herein should exhibit a selectivity ratio in
the range of about 10 to 100000. Thus, many compounds of the
invention exhibit high selectivity for inhibition of O-GlcNAcase
over .beta.-hexosaminidase.
The present invention has been described with regard to one or more
embodiments. However, it will be apparent to persons skilled in the
art that a number of variations and modifications can be made
without departing from the scope of the invention as defined in the
claims.
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